Fetal exposure to alcohol can severely and permanently damage the developing brain and can lead to fetal alcohol spectrum disorder (FASD). Despite public information campaigns and warnings from the Surgeon General, many women continue to abuse alcohol during pregnancy, and FASD remains one of the most common causes of mental retardation. In the US alone, FASD cases are estimated to cost the economy $5.4 billion annually. One of the most important ways in which alcohol disrupts brain development is by killing neurons. Loss of neurons contributes strongly to the microencephaly, behavior problems, and learning deficits in children with FASD. Utilizing pharmacological, molecular, and genetic approaches, we have discovered that a particular signaling pathway mediated by nitric oxide (NO) can protect developing neurons against alcohol toxicity. Using in vitro systems, we have found that pharmacological activation of the NO-cGMP-PKG pathway can prevent alcohol-induced neuronal death. Conversely, blockade of this pathway worsens neuronal losses. Over-expression of the gene encoding neuronal nitric oxide synthase (nNOS), which produces NO within neurons, protects neurons against alcohol-induced death. The objective of this proposal is to advance our findings beyond the in vitro setting, to determine whether the pathway can similarly protect the developing brain in vivo against alcohol toxicity and can be used as a therapeutic intervention. The first aim examines the possibility that overexpression of the nNOS gene can ameliorate alcohol- induced neuronal death and behavioral deficits. Studies in this aim will examine whether ectopically expressed nNOS in Purkinje cells, the most vulnerable neuronal population to alcohol toxicity, can protect those cells against alcohol-induced cell death and prevent alcohol-induced cerebellar dysfunction. The second aim examines the mechanism by which the NO-cGMP-PKG pathway signals its neuroprotective effects. Studies in this aim will determine whether the NO-cGMP-PKG pathway protects developing neurons against alcohol-induced death by inhibiting oxidative stress. The third specific aim examines whether the phosphodiesterase 2 inhibitor, Bay60-7550, can protect the developing brain against alcohol toxicity by activating the NO-cGMP-PKG pathway, thus inhibiting oxidative stress. Completion of this research may shed light on the role of alcohol-induced oxidative stress in FASD and may identify the NO-cGMP-PKG pathway as a promising target for preventive interventions against FASD.